Atoms and molecules with unpaired electrons possess a magnetic moment generated by the electron-spin angular momentum. The magnetic moment can be detected by electron spin resonance techniques at concentrations as small as a few picomoles. Since the electron magnetic moment is sensitive to magnetic interactions at the atomic scale, it becomes an ideal probe in a variety of research areas, from material science to structural biology. For executing experiments on paramagnetic samples by an ESR device, both an intensive microwave magnetic field and an orthogonally oriented static magnetic field at the sample position are used for exciting electron spin resonance transitions within the sample. The static magnetic field is commonly produced by a magnet system of an electron spin resonance spectrometer.
Modern ESR spectroscopy relies on a repertoire of many different continuous-wave and pulsed techniques. Among the most important ones is the pulsed electron-electron double resonance technique (PELDOR, also called DEER) which permits to measure long range distances (2-10 nm) between two paramagnetic species or spin labels in (bio)macromolecules such as nucleic acids, proteins, etc. by monitoring their dipolar interaction. The resolution of the PELDOR experiment increases at higher polarizing fields where the local symmetry of the electron wave function, which is reflected in so-called g-tensor values, is well resolved. In this case, the angular dependence of the dipolar interaction can be more accurately recorded and analysed. As an example, for spin labels based on nitroxide radicals commonly used in spin labelling of diamagnetic biomacromolecules, these fields are above 3T, corresponding to the excitation microwave frequencies above 90 GHz.
Conventionally, various structures have been established for providing monochromatic irradiation through a single mode resonator. However, to perform PELDOR experiments, two microwave frequencies are required. One frequency, called “observer” frequency, is applied to monitor a specific region of the electron paramagnetic resonance spectrum and a second frequency, called the “pump” frequency, is applied to other regions of the ESR spectrum to excite the second spin in the pair. Because two microwave frequencies are used, the bandwidth of the ESR microwave resonator is crucial. In a conventional experiment, the “observer” frequency is set to the centre of a resonance dip of the resonator. As a consequence, the “pump” frequency is placed on the side of the resonator dip, where the pumping efficiency is not as good as at the centre of the dip. On commercial W-band single mode resonators the frequency difference can be set only in the range of Δf=20-60 MHz (see Y. Polyhach et al. in “Journal of Magnetic Resonance” vol. 185, 2007, p. 118-129; and D. Goldfarb et al. in “Journal of Magnetic Resonance” vol. 194, 2008, p. 8-15). This is not sufficient to detect all possible orientations for radicals in a pair. The latter is a crucial information to study conformational changes of macromolecules, e.g. biological macromolecules (biomacromolecules). To do this one has to separate the “pump” and “observer” frequencies up to Δf=200-350 MHz. With a single mode resonator, particularly at frequencies above 90 GHz, this is impossible. A recent attempt to employ a low-Q single mode resonator for ELDOR experiments with Of up to 150 MHz (see G. Sicoli et al. “Appl. Magn. Reson.” vol. 37, 2010, p. 539-548) revealed a considerable decrease in signal sensitivity. This demonstrated that the approach of a low-Q resonator is not the right concept at high frequencies. There is a need for a dual-mode resonator, in which the frequency separation up to 350 MHz would be possible. Another point is that the frequency separation, Δf, should be tuneable.
Generally, a microwave cavity is one of the critical parts of each ESR spectrometer as the cavity influences the spectrometer performance. This is particularly true for high microwave frequencies where constraints become larger. The available microwave sources for such frequencies usually have a narrow range of frequency generation (˜400 MHz). Therefore a capability of varying the resonance frequency in a broad range is desired.
Another problem encountered is the usual need for low temperature measurements. ESR/PELDOR experiments are performed mostly at low temperatures (T˜5-70 K). These low temperatures are to be stably kept over a long time during the signal accumulation (e.g. up to 60 h), i.e. the resonator should fit into a He-flow cryostat of the ESR spectrometer, should be capable to be efficiently evacuated, and tuning should be performed from the top of the cryostat. The sample usually is inserted into the resonator with a special holder (˜1.5 m long). This complicates the design of any resonator. Furthermore, orientation of the microwave magnetic fields (B1 in ESR nomenclature) should be orthogonal to the statically applied magnetic field (B0). Finally, the quality factor, Q, of the resonator is to be efficiently high to produce strong magnetic fields at the sample position and the cavity coupling to a microwave source should be precisely adjusted. Any small perturbation of the resonator geometry reduces its quality thus reducing magnetic field intensity of the microwave mode and therefore an ESR effect.
Dual-mode microwave resonators are generally known in practice (see for example: Bruker ER4116DM dual mode resonator, Bruker BioSpin), which however are adapted for X band frequencies only (e.g. about 9 GHz) and the resonance frequencies of the modes are fixed. Furthermore, the polarization directions of the microwave fields of the two modes are perpendicular to each other. Thus, these conventional resonators are suitable for ESR measurements of symmetry-forbidden transitions only. Furthermore, various configurations of multi-mode Fabry-Perot microwave resonators are generally known (see for example: I. Tkach et al. in “Review of Scientific Instruments” vol. 75, 2004, p. 4781-4788). However, these resonators are not capable of a simultaneous excitation of a sample at different microwave frequencies.
U.S. Pat. No. 5,598,097 discloses an apparatus for ESR measurements including two or more cylindrical resonators wherein a single resonance frequency of the apparatus can be tuned by changing a distance between the cylindrical resonators.